The presence or localization of specific DNA sequences in human chromosomes can be detected in chemically fixed cells by In Situ Hybridization (ISH), a method based on complementary base-pairing between the target sequence and an oligonucleotide probe that carries a detectable tag (e.g., a fluorescent dye). However, the DNA hybridization protocol is time-consuming and the oligonucleotide probes are costly. Furthermore, ISH is normally applicable to fixed cells and is challenging to apply in live cells when desired due to the stringent conditions of hybridization which are not physiological. Live cell imaging would be required for observation of the intranuclear movements or rearrangements of a given chromosomal region that contains the array of DNA sequence(s) being targeted, and would allow investigation of how such movements or rearrangement may result in human diseases.
“TALEN” (“Transcriptional Activator-Like Effector Nuclease”) is a recently introduced method that allows specific DNA sequences to be targeted by a molecular mechanism that does not involve pairing between complementary bases in the DNA and the probe. Instead, unique arrays of amino acids are incorporated into a peptide to confer upon it a high specificity for binding to a particular DNA sequence. To date, the major application of this method has been to site-specifically direct the cutting of DNA inside cells to allow the deletion/insertion/mutation, at the cut site, of a new DNA element (“genomic engineering”). This is achieved by tethering to the peptide a DNA-cutting enzyme (the “Effector Nuclease” in the method's acronym) whose action is thus directed specifically to that DNA site.
What is need in the art are compositions and methods to detect chromosomal sites by direct binding of labeled protein sequences that are devoid of nuclease activity.